This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. E2DISP, an engineered protein domain from Geobacillus stearothermophilus pyruvate dehydrogenase subunit E2, can be used to express N-terminal fusions of heterologous peptides or proteins. E2DISP self-assembles into 24 nm virus like particles (VLPs) displaying up to 60 copies of a foreign antigen. The aims are to: (1) Design HIV-E2 Envelope (Env) fusion proteins with conserved neutralization determinants and determine their effectiveness in eliciting neutralizing antibodies (NAbs);(2) Explore immunization strategies to elicit long lasting Th1 CD4, CTL memory responses while preserving sustained antibody responses; (3) Explore optimal presentation and co-administration of E2DISP VLPs with recombinant C3d-based adjuvants;and (4) Optimize the use of pure and hybrid E2-HIV VLPs in prime-boost regimens with other vaccine delivery systems effective in boosting cellular immunity and NAb responses. In the last year, we have developed effective methods for purifying and stabilizing pure and hybrid particles and have shown that mice immunized with the Gag-E2 and Env-E2 VLPs elicit CD8+ T cell responses. In experiments in rabbits using HIV Env fusion protein 60-mer particles, we have been successful in showing the development of high titers of neutralizing antibodies within only six weeks, after two immunizations with DNA and E2 protein VLPs. Both the V3 region as well as portions of the gp41 protein elicit neutralizing antibodies in rabbits, and we are exploring the breadth of this response. We are also exploring the delivery of these fusion proteins with microneedle delivery to the mucosal antigen-presenting cells.